Summary
Five mycoplasma species most frequently isolated from cell cultures were tested for the presence of endogenous hypoxanthine phosphoribosyl-transferase (HPRT), activity. All of the five, cultured in cell-free medium, contained variable but significant levels of HPRT. Two strains ofM. hyorhinis exhibited a 13-fold difference in their specific HPRT activity. When infected with any of these mycoplasma species, HPRT-deficient mouse cell mutants rapidly acquired a cell-associated HPRT activity; however, the cells remained sensitive to HAT medium and resistant to 6-thioguanine. On the other hand, normal HPRT-positive cells deliberately infected with the mycoplasmas uniformly became sensitive to HAT medium. The apparent transfer of mycoplasma-specific HPRT activity to HPRT-deficient cells may be used as a sensitive measure of cell infection by these mycoplasma strains. The HPRT activities of mycoplasmas share several common properties so that they can be distinguished easily from the mammalian HPRT isozymes. Compared to the animal cell enzymes, the mycoplasmal HPRT activities are less heat stable, more strongly inhibited by 6-thioguanine, and in general migrate more slowly in electrophoresis at a neutral pH.
Similar content being viewed by others
References
Stanbridge, E. 1971. Mycoplasmas and cell cultures. Bacteriol. Rev. 35:206–227.
McGarrity, G. J. 1975. Detection of mycoplasma in cell cultures. Procedure 75317. TCA Manual 1: 113–116.
Schneider, E. L., E. J. Stanbridge, and C. J. Epstein. 1976. Incorporation of3H-uridine and3H-uracil into RNA. Exp. Cell Res. 84:311–318.
Levine, E. M. 1974. A simplified method for the detection of mycoplasma. Methods Cell Biol. 18: 229–248.
Chen, T. R. 1977.In situ detection of mycoplasma contamination in cell cultures by fluorescent Hoechst 33258 stain. Exp. Cell Res. 104: 255–262.
Van Diggelen, O. P., D. M. Phillips, and S. Shin. 1977. Endogenous HPRT activity in a cryptic strain of mycoplasma and its effects on cellular resistance to selective media in infected cell lines. Exp. Cell Res. 106:191–203.
Barile, M. F., H. E. Hopps, M. W. Grabowski, D. B. Riggs, and R. A. DelGiudice. 1973. The identification and sources of mycoplasmas isolated from contaminated cell cultures. Ann. N.Y. Acad. Sci. 225: 251–264.
Stanbridge, E. J., J. A. Tischfield, and E. L. Schneider. 1975. Appearance of hypoxanthine guanine phosphoribosyltransferase activity as a consequence of mycoplasma contamination. Nature 256: 329–331.
Van Diggelen, O. P., S. Shin, and D. M. Phillips. 1977. Reduction in cellular tumorigenicity after mycoplasma infection and elimination of mycoplasma from infected cultures by passage in nude mice. Cancer Res. 37: 2680–2687.
Earle, W. R. 1943. Production of malignancy in vitro. IV. The mouse fibroblast cultures and changes seen in the living cells. J. Nat. Cancer Inst. 4: 165–212.
Littlefield, J. W. 1964. Three degrees of guanylic acid-inosinic acid pyrophosphorylase activity-mouse fibroblasts. Nature 203: 1142–1144.
McGarrity, G. J., and L. L. Coriell. 1973. Detection of anaerobic mycoplasmas in cell cultures In Vitro 9: 17–18.
DelGiudice, R. A., N. F. Robbilard, and T. R. Carski. 1967. Immunofluorescence identification of mycoplasma on agar by use of incident illumination. J. Bacteriol. 93: 1205–1209.
Clyde, W. A. 1964. Mycoplasma species identification based on growth inhibition by specific antisera. J. Immunol. 92: 958–965.
Van Diggelen, O. P., and S. Shin. 1974. A rapid fluorescence technique for electrophoretic identification of hypoxanthine phosphoribosyltransferase allozymes. Biochem. Genet. 12: 375–384.
Shin, S. 1974. Nature of mutations of conferring resistance to 8-azaguanine in mouse cell lines. J. Cell Sci. 14: 235–251.
Levine, E. M. 1972. Mycoplasma contamination of animal cell cultures: A simple, rapid detection method. Exp. Cell Res. 74: 99–109
Perez, A. G., J. H. Kim, A. S. Gelbard, and B. Djordjevic. 1972. Altered incorporation of nucleic acid precursors by mycoplasma-infected mammalian cells in culture. Exp. Cell Res. 70: 301–310.
Holland, J. F., R. Korn, J. O'Malley, H. J. Minnemayer, and H. Tieckelman. 1967. 5-Allyl-2′-deoxyuridine inhibition of nucleoside phosphorylase in HeLa cells containing mycoplasma. Cancer Res. 27: 1867–1873.
Author information
Authors and Affiliations
Additional information
This work was supported in part by PHS Research Grants 5 R01 GM21014 and 1 P03 GM19100 (Genetics Center Grant to Albert Einstein College of Medicine), and PHS Research Contracts N01 GM 6-2119 and N01-AG-4-2865 (to the Institute for Medical Research), from the National Institute of General Medical Sciences and National Institute on Aging. S. S. is a recipient of a Faculty Research Award from the American Cancer Society.
Rights and permissions
About this article
Cite this article
van Diggelen, O.P., McGarrity, G.J. & Shin, SI. Endogenous HPRT activity in mycoplasmas isolated from cell cultures. In Vitro 14, 734–739 (1978). https://doi.org/10.1007/BF02617965
Issue Date:
DOI: https://doi.org/10.1007/BF02617965